Mast cells are the major effector cells in diseases of immediate hypersensitivity. Understanding how mast cells develop from primitive blood cells may have implication for treatment of certain allergic diseases. Because mast cells may be uniquely related to hematopoietic stem cells, studies of early events in mast cell differentiation may have broader implications for hematopoiesis in general. The research proposed in this grant application extends the mast cell model system developed in the previous grant period. This system takes advantage of the fact that committed progenitors, but not uncommitted ones, can respond to stem cell factor alone by production of mast cell colonies in methylcellulose. There is continuing attention to the different roles that the c-kit/stem cell factor receptor-ligand complex may play in different stages of differentiation may have broader implications for hematopoiesis in general. The research proposed in this grant application extends the mast cell model system developed in the previous grant period. This system takes advantage of the fact that committed progenitors, but not uncommitted ones, can respond to stem cell factor alone by production of mast cell colonies in methylcellulose. There is continuing attention to the different roles that the c-kit/stem cell factor receptor-ligand complex may play in different stages of differentiation; however, the major focus in this grant application addresses Fc receptors on mast cells and their progenitors, because of recent provocative findings that in mast cells, both FcRI and FcRIII associated gamma chain dimers to transduce signals for activation and cytokine biosynthesis. We are particularly interested in the possibility that an induced autocrine IL-3 loop may be critical element in the committed state of these progenitors. The first specific aim is to determine whether bone marrow progenitors, particularly those which may be part of the early mast cell lineage, bear FcRI and FcRIII and if these receptors transduce a signal to induce an autocrine IL-3 loop which is necessary for commitment. In the second specific aim, mast cell-committed progenitors or granulated mast cells from MLN or peritoneal washouts will be tested to determine if FcRI/FcRIII-mediated signalling affects expression of autocrine cytokines, GATA transcription factors, or proteases. We will also determine if the continued occupancy of FcRI on mast cells in necessary for continued division of successive generations of daughter cells. The third specific aims takes advantage of an exciting new models system for the study of hematopoiesis; embryonic stem (ES) cell cultures. Once these very primitive cells are allowed to differentiate in culture, the expression of message and protein for FcRI/FcRIII will be monitored to correlate with the appearance of mast cells and autocrine IL-3 inducibility. These experiments may reveal similarities or differences between proposed in this grant application should contribute not only to our understanding of mast cell differentiation but also to Fc receptor ontogeny and stem cell development.
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